We propose to perform a genetic and biochemical analysis of the mechanisms of osmotic regulation in the bacterium Salmonella typhimurium. The project is based on the observations that exogenous proline is stimulatory to S. typhimurium during osmotic stress, and that osmotic stress enhances the activity of two proline permeases, the ProP and the ProU systems. We shall investigate the mechanism of osmoregulatin in S. typhimurium by these specific aims: 1) we shall determine the effect of osmotic stress on the kinetic parameters of the ProP system, 2) we shall determine the effect of mutations in various proline transport systems on the intracellular proline levels, 3) we shall isolate pleiotropic osmoregulatory mutations that alter the expression of the proU gene and other genes subject to osmotic regulation and 4) we shall isolate, clone, and sequence mutations in the promoter region of the proU gene. The results obtained in the study of osmoregulation in bacteria are of value not only from the standpoint of basic science, but also because they have potential applications to the understanding of osmoregulation in plants and the genetic engineering of plants with increased tolerance of osmotic stress.